Age-related heart valve disease is the 3rd leading cause of cardiovascular disease and is especially prevalent among the elderly. Studies have shown that degenerative aortic valve disease affects over 25% of people over 65 years of age, leading to calcific aortic valve disease (CAVD) in 4-8 years. Currently, the only effective long- term treatment for advanced CAVD is valve replacement surgery, an invasive, high-risk procedure for elderly patients. For this reason, a non-invasive therapeutic to stop the progression of CAVD would greatly benefit those most at risk for developing severe CAVD. At the cellular level, CAVD is believed to be initiated by activation of AV interstitial cells (AVICs)to a myofibroblast phenotype. Once activated, these cells increase extracellular matrix deposition, particularly type I collagen, which directly leads to decreased compliance of the leaflets observed in CAVD. Transforming growth factor-b1 (TGF-b1) has been the most extensively studied cytokine initiator of CAVD; however, serotonin (5-HT) and other serotonergic receptor agonists have been shown to lead to CAVD upstream of TGF-b1 through signaling at the serotonergic receptor, 5-HT2B. Genetically, NOTCH1 haploinsufficiency results in CAVD with 100% penetrance in human patients. Notch1+/- leads to increased synthesis and/or signaling of both BMP2 and TGF-b1 in the AV leaflets of mice, which results in CAVD. Therefore, Notch1+/- mice provide a clinically relevant animal model to examine strategies against CAVD. Here, we show evidence that 5-HT2B antagonism prevents phenotypic alteration of AVICs by TGF-b1 in vitro. Moreover, we present evidence that 5-HT2B antagonism prevents non-canonical TGF-b1 signaling in AVICs. We therefore hypothesize that 5-HT2B can be specifically antagonized to prevent AVIC myofibroblast activation and provide early time point molecular targets to treat CAVD and we will test this hypothesis in Notch1+/- animals and isolated AVICs both for these animals and humans. We anticipate that this research plan will demonstrate a novel treatment strategy for CAVD. Further, this research plan will elucidate the underlying molecular mechanism, while also quantifying biomechanical changes and functional hemodynamics due to 5-HT2B antagonism.